Phage antibodies for the immunochemical characterization of Herbaspirillum seropedicae Z78 glycopolymers

Abstract

Purpose Microbial carbohydrate antigens are targets of the immune systems of hosts. In this context, it is of interest to obtain data that will permit judgment of the degree of heterogeneity, chemical makeup, and localization of the antigenic determinants of the Herbaspirillum surface glycopolymers. Methods A sheep single-chain antibody-fragment phage library (Griffin.1, UK) was used to obtain miniantibodies to the exopolysaccharides (EPS-I and EPS-II), capsular polysaccharides (CPS-I and CPS-II) and lipopolysaccharide (LPS) of Herbaspirillum seropedicae Z78. To infer about the presence or absence of common antigenic determinants in the cell-surface polysaccharides of H. seropedicae Z78, we ran a comparative immunoassay using rabbit polyclonal and phage recombinant antibodies to the surface glycopolymers of H. seropedicae Z78. Results We isolated and purified the exopolysaccharides (EPS-I and EPS-II), capsular polysaccharides (CPS-I and CPS-II), and lipopolysaccharide (LPS) of Herbaspirillum seropedicae Z78. Using rabbit polyclonal antibodies, we found that these cellsurface polysaccharides were of a complex nature. EPS-I, EPS-II, CPS-I, CPS-II, and LPS contained common antigenic determinants. CPS-I, CPS-II, and LPS also contained individual antigenic determinants composed of rhamnose, N-acetyl-D-glucosamine, and N-acetyl-D-galactosamine-sugars responsible for cross-reactions with miniantibodies. Conclusions The anti-LPS miniantibodies were more specific for the core region of the LPS, in which rhamnose was the most abundant sugar, than they were specific for its O portion. The miniantibodies we isolated can be useful reagents not only in basic biochemical research but also in clinical diagnostic and therapeutic applications.